Apparatus for macromolecule purification

ABSTRACT

An electrophoresis apparatus, comprises a cartridge configured to be removably mounted in the apparatus, and a separation membrane positioned in the cartridge. The separation membrane has a first side along which a first flow path defined in a first grid element and a second side along which a second flow path defined in a second grid element is provided. Restriction membranes separate buffer flow from the first flow path and the second flow path. The apparatus includes connection blocks which house electrodes and inlets and outlets for buffer flow and sample flows. The cartridge is removable from the connection blocks for replacement with another cartridge.

CROSS REFERENCE TO PARENT APPLICATION

This application is a Continuation Application of U.S. patentapplication having Ser. No. 09/390,565 filed on Sep. 3, 1999, now U.S.Pat. No. 6,328,869.

FIELD OF THE INVENTION

This invention relates to an apparatus for purification ofmacromolecules in solution and in particular to a cartridge or cassettefor use in a macromolecular purification apparatus.

BACKGROUND OF THE INVENTION

European patent No. 352286 relates to improvements in the separation ofmacromolecular solutes by a process known as electrophoretic separation,in particular fixed boundary electrophoretic separation. In fixedboundary electrophoresis, as is explained in more detail below in thedescription following the Brief Description of the Drawings, asemi-permeable membrane (hereinafter referred to as a separationmembrane) acts to separate two streams of liquid carrying macromolecularsolutes such as proteins, referred to as the upstream and thedownstream. The streams pass between charged electrodes and at least onemacromolecular solute migrates across the membrane from one stream tothe other under the influence of the electric field. The apparatus alsoincludes flow paths for buffer solution and further semi-permeablemembranes, hereinafter referred to as restriction membranes, disposedbetween the electrodes and the separation membrane. The restrictionmembranes allow the passage of ions but not macromolecules.

The present invention relates to developments and improvements over theideas and principals disclosed in EP 352286 and in particular torefinements and improvements to the apparatus to make the technologyeasier to use and operate.

In particular, it is one desired object of the present invention toprovide an apparatus which is easier to fill, empty, clean andreassemble in contrast to existing apparatus such as that describedabove which tends to be formed substantially integrally with storagetanks, pumps, cooling apparatus and similar features making theapparatus expensive and difficult to handle and clean.

The lack of ease of use of the prior art apparatus is a seriousdisadvantage and tends to make separation of molecules byelectrophoresis an unnecessarily complicated, tedious, time-consumingand expensive procedure.

Thus it is a further object of the present invention to provide anelectrophoresis apparatus which is simpler and relatively quicker tooperate than existing apparatus to set up and use.

A further problem associated with the prior art id the amount of“downtime” involved after the apparatus has been used and before it canbe set up for a further separation. Electrophoresis apparatus isexpensive and the downtime is consequently a serious cost disadvantage.

Thus it is a further object of the present invention to provide anapparatus whose downtime is reduced in comparison with prior artapparatus.

A yet further problem of existing electrophoresis apparatus relates tothe size of the samples which are typically separated by such apparatus.The amount of sample to be separated can be very expensive, andconsequently the smaller the sample which can be separated by theapparatus, the better.

It is a yet further object of the present invention to provide anapparatus which can be used to separate relatively small samples sizes.

Existing electrophoresis apparatus is also very bulky and can take up alarge area of laboratory space.

It is a yet further object of the present invention to provide anapparatus which in its preferred embodiment can be relatively compact.

A further problem with existing electrophoresis apparatus is that it canrequire a relatively high current and voltage to operate.

It is a further object of the present invention to enable anelectrophoresis apparatus which uses less power and hence is moreeconomical and also causes less electrical heating to the sample beingseparated which can damage the sample.

SUMMARY OF THE INVENTION

In a first broad aspect, the present invention provides an apparatus forfree flow electrophoresis in which a separation membrane, a first flowpath along one side of the separation membrane, and a second flow pathalong an opposite side of the separation membrane and restrictionmembranes for separating buffer flow from the flow paths, are housed ina cartridge which can be removed from the apparatus for replacement withanother cartridge, after use.

The construction of an apparatus in which the separation membrane andflow paths are defined in a removable cartridge provides a modularsystem which has substantial advantages of the known prior art. The needto clean, readjust, and re-set the apparatus after a separation has beencarried out is reduced. The used cartridge is easily removed and a newcartridge may be simply inserted. The upstream and downstream circuitsonly require cleaning although the buffer may be replaced if desired.

Thus the present invention advantageously provides an apparatus whosedowntime is greatly reduced in comparison with prior art apparatus. Thisproduces substantial savings in terms of time and cost efficiency andmay also enable the separation of samples at a more commercial scalethan is possible with existing electrophoresis apparatus.

In one particular preferred embodiment, the apparatus includes an upperand a lower connection block which defines inlet and outlet means forfeeding liquid into the first flow path (the downstream) and into thesecond flow path (the upstream), in the cartridge.

This arrangement enable the cartridges to be simply inserted between thetwo blocks and the blocks may be brought together, by a screw clamp orthe like to engage the cartridge.

The upper and lower connection blocks may house electrodes andconnection means for connecting the electrodes to a power source.

In one particularly preferred embodiment, the electrodes are made oftitanium mesh coated with platinum.

The standard electrodes usually used for prior art electrolytic cellscomprise platinum wire. The platinum coated titanium expanded mesh usedin accordance with preferred embodiments of the present invention hasseveral advantages over platinum wire in particular the ridged structureis self supporting and less expensive than platinum wire. The mesh alsomay provide a greater surface area and tends to allow higher currents topass through the separation unit with less electrical power losses ofthe electrode. The electrodes may thus have a longer useful life.

The upper and lower connection blocks may be made of any suitablematerial for example a plastics material. The blocks may be transparentor opaque.

The upper and lower electrodes may be housed in recesses or channelsdefined in the upper and lower connection blocks, respectively. Thosechannels may define part of the boundaries of the buffer flow path forthe apparatus. The other boundary for the buffer flow path is defined bythe restriction membranes housed in the cartridge which form the upperand lower faces of the cartridge. The upper and lower connection blocksmay define inlets and outlets for buffer flow.

In a related aspect the invention also encompasses a cartridge for usein the apparatus of the present invention, the cartridge including ahousing and containing a separation membrane, a first flow path definedalong one side of the separation membrane, and a second flow pathdefined along an opposite side of the separation membrane andrestriction membranes for isolating buffer flow from the first andsecond flow paths.

The cartridge may further include gaskets positioned either side of therestriction membranes for sealing the components between the upper andlower connection blocks. The first and second flow paths may be definedby first and second grid elements. Each grid element may be generallyplanar and relatively thin.

The use of a relatively thin planar grid element defining the first andsecond flow paths may provide substantial advantages over the existingart. First, a relatively thin grid element tends to increase the liquidvelocity and pressure and thus may result in a more even distribution ofliquid over the separation membrane in the cartridge. The resultantrelatively higher liquid velocity may also assist in inhibiting foulingof the separation membrane.

Also, the volume of liquid required may be decreased by the use of arelatively thin grid which enables relatively smaller sample volumes tobe used for laboratory separations and this may be a significantadvantage over the prior art when the samples are expensive to produce.

Finally, the use of a relatively thinner grid element may enable lesselectrical power to be deposited into the liquid since power supplied tothe liquid across the grid region, depends on the thickness of the grid.Thus, a relatively smaller power supply unit may be used, and this maymake the process more economical. If less heat is transferred into theliquid the temperature of the liquid remains lower. This is advantageoussince high temperatures may destroy the sample and product.

The grid element and restriction membranes may include a through holelocated near each end of the element/membrane for the passage ofupstream and downstream liquid therethrough to the grid element. Thegrid element may include a central elongate at least part cut outportion which in the assembled cartridge in conjunction with the twoadjacent membranes, defines a flow path or channel from the hole one endof the grid element to the hole at the other end. The flow path mayinclude a lattice for supporting the separation membrane and for mixingthe flow.

All the elements of the cartridge apart form the cartridge housing orcasing are generally planar and are assembled together in a sandwichtype construction.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention will now be described, by way of example only, and withreference to the accompanying drawings in which:

FIG. 1 schematically illustrates an example of charge based separation;

FIG. 2 schematically illustrates an example of size based separation;

FIG. 3 illustrates an example of concentration;

FIG. 4 illustrates an example of dialysis;

FIG. 5 is a schematic diagram of a separation unit in accordance with anembodiment of the present invention;

FIG. 6 is a schematic diagram of an apparatus in accordance with anembodiment of the present invention which includes the separation unitof FIG. 5;

FIG. 7 is a section through a clamp means which forms part of anapparatus in accordance with an embodiment of the present invention;

FIG. 8 is a exploded view of a cartridge which may be used with anembodiment of the present invention;

FIG. 9A is a plan view of a grid element which may be a component of thecartridge of FIG. 8;

FIG. 9B is a reverse plan view of the grid element of FIG. 9A;

FIG. 10 is a cross-section on lines X—X of FIG. 9A;

FIG. 11 is a cross-section on lines XI—XI of FIG. 9A;

FIG. 12 is a cross-section on lines XII—XII of FIG. 9A; and

FIG. 13 is a plan view of a grid element which may be used with a secondembodiment of the present invention.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

Before describing a preferred embodiment in detail, some of theprincipals of operation of an apparatus in accordance with the presentinvention will first be described. The description is not intended tolimit the present invention to any one particular principal or theory ofoperation.

An electric field applied to macromolecules, such as proteins, insolution, will tend to cause the proteins to move to the electrodes. Ifthe protein has a positive charge, it will tend to move to the negativeelectrode (cathode). Conversely, a negatively charged protein will tendto move to the positive electrode (anode).

In an apparatus in accordance with the present invention, a separationmembrane may be placed in the electric field and molecules may beselectively transported between two circulating streams which may becalled the upstream and the downstream. The particular separationmembrane used may vary for different applications and may generally havea relatively large, but relatively well defined, pore size. The upstreamand downstream may be isolated from the electrodes by buffer streams andby two restriction membranes. The restriction membranes may allow themovement of small molecules and ions up to a molecular weight of about3,000 daltons.

An apparatus in accordance with the present invention may be operated inat least four different modes. Examples of those modes are set out belowand are illustrated in FIGS. 1 to 4, respectively.

1. Charge Based Separation

In principal, any two molecules with different pIs may be separated bycarrying the separation out at a pH between the two pIs. The pI is thepH of a solution in which the molecule has neutral charge, thus bychanging the pH of the solution in which a molecule is present, theeffective charge of that molecule may be changed.

Thus in a solution with a pH between the two pIs, one molecule will havea positive charge, and will tend to move towards the cathode and tend tobe contained in the upstream. The other molecule will have a negativecharge and will tend to be contained in the downstream as it will tendto move towards the anode. FIG. 1 illustrates an example of charge basedseparation.

2. Sized Based Separation

Components with different molecular weights may be separated on thebasis of pore size. FIG. 2 shows a mode of separation based on size. Twoproteins which are both negatively charged, may be separated because thelarger molecules tend to be unable to migrate through the smaller poresof the separation membrane. Careful combination of pore size and pH mayoften allow the isolation of a single component from a complex mixturein one run. (Examples are Mab from ascitic fluid and ovalbumin,lysozyme, or avidin from egg white, and fibrinogen from plasma).

3. Concentration

FIG. 3 illustrates an example of concentration, which tends to utilize arelatively large pore size separation membrane (1,000 kDa). Therelatively large pore sizes may enable the rapid transportation ofproteins across the separation membrane from a large volume upstreamsolution to a small volume downstream solution. In this process a pH maybe selected in which all of the desired proteins will tend to have thesame charge. Typically, pH 8.3 is selected since most proteins will havea negative charge at this pH. In many applications, purification may beachieved at the same time as concentration.

4. Dialysis

FIG. 4 illustrates an example of dialysis. The apparatus in accordancewith the present invention may be operated for desalting. The separationmembrane is not necessary for dialysis, which can be performed with orwithout this membrane. The ions may be removed from the sample bypassing through the restriction membranes and they then tend to bewashed away by the outer buffer streams. This process may also occurduring standard fractionation or concentration, but in these cases theions are usually recirculated. When being used for extensive dialysis,the outer buffer, where the ions collect, should preferably be exchangedfor fresh buffer solution at regular intervals.

Having outlined some of the principals of operation of an apparatus inaccordance with the present invention, we now turn to a description ofthe apparatus itself with reference to FIG. 5 which is a schematiccross-section through an example of a separation unit in accordance withthe present invention.

The separation unit may comprise an electrolytic cell which may includefour chambers. The two outer chambers 12 and 14 may house the electrodesand a flow of buffer may pass through these chambers. The inner chambersmay be called the upstream and downstream channels which may beseparated from the outer buffer chambers by restriction membranes 20 a,20 b. A separation membrane 22 may separate the upstream and downstreamchambers. The upper negative electrode 24 may be referred to as thecathode and the lower positive electrode 26 maybe referred to as theanode.

A schematic diagram of an exemplary embodiment of an apparatus inaccordance with the present invention is shown in FIG. 6. As shown inthe illustrative example, the four chambers 12 to 18 are connected tothree flow circuits. Each flow circuit may consist of a reservoir and acirculating pump. Thus the upstream reservoir may comprise upstreamreservoir 30 and pump 32 and associated tubing, the downstream circuitmay comprise downstream reservoir 34, pump 36 and associated tubing andthe buffer circuits may share a single buffer reservoir 38 and a singlebuffer pump 40. The circuit may divide into two paths prior to passingthrough the two buffer chambers 12 and 14 and after passing through thechambers, the two paths may rejoin.

The separation unit may be cooled by various means such as ice bricks orcooling coils (external apparatus) placed in the buffer reservoir tocontrol its temperature, or by any other suitable cooling means. Theupstream and downstream reservoir flow paths tend to pass through thebuffer reservoir and heat may be exchanged between the upstream and thebuffer reservoir, and the downstream and the buffer reservoir. This heatexchange tends to maintain a low temperature in the upstream anddownstream, which is preferred for protein separation.

In the purely illustrative embodiment described above, the buffer pump40 may be incorporated into the apparatus. The pumps 32 and 36 for theupstream and downstream may be external or internal.

The electrodes may be connected to an external power supply 50 viaelectrical connecting leads. The power supply may ideally have avariable voltage up to 200 V and may be capable of delivering up to oneamp. Relatively higher voltages, up to 500 V may be used provided thatthe total power (voltage x current) is maintained at less than about 200W.

The buffer solution provides the ions which may provide the current flowin the electrolytic cell, The buffer also tends to stabilise the pHduring separation and to act as the cooling medium. The buffer flow mayalso help prevent build up of gases at the electrodes generated byelectrolysis.

FIG. 7 shows an example of a separation unit housing a cartridge forcassette) and FIG. 8 shows an exploded view of the cartridge of FIG. 7.The separation unit may include an upper connection block 102 and alower connection block 104 between which, in use, the cartridge may beclamped.

The cartridge may comprise a cartridge housing 202 which may hold 10 thecomponents of the cartridge. The cartridge may be generally elongate andmay include two parallel elongate side walls 203 which may extend alongthe longitudinal axis A—A of the cartridge. Each end of the cartridgemay include three end walls so that the cartridge may be generallyoctagonal in plan view. A small flange 206 may extend around the base ofthe walls 203, 204. The flange may project inwards towards the centre ofthe cartridge. On the right hand side of the cartridge, as oriented inFIG. 8 a flap 208 may be provided which may act as a handle for holdingor manipulating the cartridge. A planar silicon rubber gasket 210 whoseexterior is generally octagonal may be configured to fit inside thewalls on the cartridge resting on the flange 206. The centre of thegasket may define an elongate cut out portion 212. Adjacent either endof the seal a small cylindrical hole 214, 216 may be provided.

Above the gasket 210 a first restriction membrane 20 b may be located.The external shape of the gasket may be generally the same as that ofthe interior of the cartridge so that it too may fit inside thecartridge. The membrane may have two holes 218, 220 adjacent either endof the membrane and positioned so that when the cartridge is assembled,those holes tend to align with the holes 214, 216 of the gasket 210. Therestriction membrane may typically be formed from a thin substrate andcoated with compounds to give a relatively small pore size, such aspolyacrylamide gel, so that only very small particles such as ions tendto pass through the gel. such pores tend to be too small to allow thepassage of macromolecules such as proteins.

Above the restriction membrane there may be a first grid element 230.Above that grid element 230 the separation membrane 22 is located. Thepore size of the separation membrane used may depend on the size of theparticular molecules being separated and the actual mode of separationbeing used.

One function of the grid element 230 is to tend to keep membranes 20 aand 22 apart. The grid element also may provide a flow path for theupstream (or the downstream, since the grid elements for both may beidentical). The grid element may be generally planar and the exterior ofthe grid element may be shaped to fit inside the walls of the cartridgehousing. An example of a grid element in accordance with the presentinvention is shown in more detail in FIGS. 9 through 12. An elongaterectangular cut-out portion 231 which incorporates lattice work may bedefined in the centre of the grid element. At each end of the grid athrough hole 232 may be provided, which in the assembled cartridge, maybe aligned with holes 214 and 218 of the gasket and restrictionmembrane, respectively. A triangular channel area 234 having sides and abase, may extend and diverge from the hole to the cut out portion 231.Upstanding ribs 236, 238, 240, best seen in FIGS. 11 and 12, may bedefined in the channel area 234. Liquid flowing up through the hole 232may thus pass along the triangular channel area 234 between the ribs andinto the lattice. The ribs may direct the flow from the hole 232 andtend to ensure that the liquid is evenly distributed along thecross-section of the lattice. The ribs may also provide support to theseparation membrane 22 disposed above the grid element. The narrowchannels defined by the ribs also tend to increase the liquid velocityand pressure in the downstream.

The lattice may comprises a first array of spaced parallel members 24125 extending at 450 to the longitudinal axis of the grid disposed aboveand integrally formed with a second lower set of spaced parallel members242 extending at 900 to the first set of members. As illustrated in theexample shown in FIG. 10, the upper and lower surfaces of the membersmay be rounded. The absence of any sharp edges helps prevent damage tothe separation membrane 22. Further, any increased pressure on theseparation membrane 22 will tend to increase the area of contact betweenthe curved lattice members and the separation membrane and may thusprovide extra support to the separation membrane. The lattice may evenlydistribute the flow of liquid over the separation membrane surface. Theuse of a first set of members disposed above a second set of memberstends to ensure that the liquid in the downstream is forced to move upand down and change direction frequently and this helps to encouragemixing of the liquid and tends to inhibit static flow zones.

The reverse side of the grid element as illustrated in figure SB may berelatively smooth and flat aside from the cut-out 231 and the holes 232,which tends to ensure sealing between the buffer stream and thedownstream.

The thickness of the grid element may be relatively small. Withreference to FIG. 10, the exterior areas of the element may be 0.8 mmthick. A sealing rib or ridge 252 may extend around the periphery of thegrid to improve sealing. The ridge may be 1.2 mm thick measured from oneside of the grid element to the other. The distance between oppositepeaks of the lattice elements 241, 242 measured from one side of thegrid to the other may be 1 mm. The relative thinness of the grid mayprovide several advantages. First, a relatively thin grid element tendsto increase the liquid velocity and pressure and thus may result in amore even distribution of liquid over the separation membrane 22. Theresultant relatively higher liquid velocity may also assist ininhibiting fouling of the separation membrane 22. Fouling may occur whenmacromolecules block the pores of the separation membrane or stick tothe separation membrane.

Also, the volume of liquid required may be decreased by the use of arelatively thin grid which enables relatively smaller sample volumes tobe used for laboratory separations and this may be a significantadvantage over the prior art when the samples are expensive to produce.

Finally, the use of a relatively thinner grid element may enable less 25electrical power to be deposited into the liquid since power supplied tothe liquid across the grid region, depends on the thickness of the grid.Thus, a relatively smaller power supply unit may be used, and this maymake the process more economical. If less heat is transferred into theliquid the temperature of the liquid remains lower. This is advantageoussince high temperatures may destroy the sample and product.

The separation membrane 22 may be selected depending on the applicationas discussed earlier in the description relating to FIGS. 1 to 4 of thedrawings. Following the separation membrane there may be a mirror imageof elements 210, 220: there is a further grid element 230, a restrictionmembrane 20 a, and a further gasket 210 which may be symmetricallyarranged about the separation membrane 22. Those three components mayform the upstream and a part of the boundary of the buffer stream. Thecomponents may be held in the cartridge 200 by means of a clip 260 whichmay be snap fitted or glued around the top of the walls 204 of thecartridge.

One function of the cartridge is to hold the components together for 5insertion into the separation unit. The actual cartridge walls tend tohave no effect on the sealing of the system. In accordance with oneembodiment of the invention the system may be sealed in manner in whichno liquid contacts the walls of the cartridge in use.

FIG. 7 illustrates an example of a cartridge located in the clamping 10unit between connecting blocks or jaws 102 and 104. The jaws may housethe inlets and outlets for feeding buffer flows, a downstream and anupstream into the cartridge. The jaws may include inlets 304 and 306which respectively may allow the upstream and downstream flows into thegrid via the holes 214, 216, 230. Outlets 308 and 310 may be similarlyprovided. Inlets 312 and 314 and outlets 316 and 318 may feed bufferpast the restriction membranes 20 a, 20 b along the flow path definedbetween a recess or channel 320 in the lower jaw 104 in which the anodemay be located, and the gasket 210 and the restriction membrane 20 b. Asimilar arrangement may be provided in the upper jaw which may house thecathode. The jaws may be mounted between a simple screw clamp unit sothat a simple screw operated knob may be used to open and close the jawsfor changing the membrane cartridge.

The electrodes 26, 24 may be formed from platinum coated titaniumexpanded mesh, in contrast with the standard electrodes usually used forelectrolytic cells which comprise platinum wire. The platinum coatedtitanium expanded mesh used in the apparatus of the present inventionhas several advantages over platinum wire in particular the ridgedstructure is self supporting and less expensive than platinum wire. Themesh also may provide a greater surface area and tends to allow highercurrents to pass through the separation unit with less electrical powerlosses of the electrode. The electrodes may thus have a longer usefullife.

Finally, the larger surface area distributed over the buffer channel mayprovide a more even electrical field for the separation process.

The electrodes may also be located relatively close to the restriction35 membranes. This tends to enable less electrical power to be used anddeposited into the buffer liquid and consequently less heating of thebuffer liquid may occur. Connectors 300 and 302 from the electrodes maypass to sockets for connection of electrical power to the electrodes.The electrodes may be shrouded to prevent accidental contact with anoperators fingers or the like.

In use, a cartridge in accordance with the present invention may beloaded into the apparatus, the jaws may be closed to seal the componentsin place, the buffer solution and upstream and downstream may be fedthrough the connection blocks and thence through the cartridge and theapparatus may be connected to a power supply. The electrical power maybe set to the desired value and separation may be carried out using oneof the modes described earlier with reference to FIGS. 1 to 4. After theseparation has been carried out, the used cartridge may simply beremoved and replaced with another cartridge ready for re-use. Theupstream and downstream sample circuits may be cleaned and the bufferreplaced if necessary. Following that the apparatus is ready to carryout a further separation.

FIG. 13 illustrates a grid element 270 for a second embodiment of thepresent invention which utilises a separation membrane having a muchlarger surface area than that of grid element 230. The principal ofoperation of this grid element may be generally the same as that of thesmaller grid element 230 although the holes 272 through which thedownstream or upstream is fed are located in two of the corners of thegrid element and there may be more channels 273 feeding the streams fromthe holes 272 to the central portion 274 of the grid. The cartridge,casing and other components of the cartridge may also be relativelyincreased in area so that they have approximately the same area as thegrid.

1. A removable cartridge for use in an electrophoresis unit comprising:a housing having at least one sidewall sealingly connected theretodefining an interior; a first restriction membrane disposed within theinterior; a second restriction membrane disposed within the interior andgenerally in parallel with the first restriction membrane; a separationmembrane disposed in the interior between the restriction membranes; afirst spacer having a grid element disposed between the firstrestriction membrane and the separation membrane so as to define a firstseparation chamber therebetween; a second spacer having a grid elementdisposed between the second restriction membrane and the separationmembrane so as to define a second separation chamber therebetween; meansdefining a first fluid pathway through the first separation chamber;means defining a second fluid pathway through the second separationchamber; and at least one gasket disposed within the interior of thehousing and proximate to an outer surface of a selected one of therestriction membranes so as to be adapted to sealingly engage anassociated fluid pathway to allow for an electrically conductive buffersolution to flow along the at least one restriction membrane; whereinthe cartridge is adapted to be removably engaged in an electrophoresisseparation unit such that in use selected compounds in the first orsecond fluid pathway are caused to move through one or more of themembranes under the influence of an electric field.
 2. The cartridgeaccording to claim 1 further comprising: first inlet means adapted forcommunicating a first associated fluid into the first separationchamber; second inlet means adapted for communicating a secondassociated fluid into the second separation chamber; first outlet meansfor outputting fluid from the first separation chamber; second outletmeans for outputting fluid from the second separation chamber; such thatin use the first and second associated fluids stream through theseparation chambers without substantial convective mixing of fluidsbetween the chambers.
 3. The cartridge according to claim 2 wherein themembranes are comprised of a material adapted to allow for passage of anelectric field therethrough such that a portion of compounds in thefluids are caused to move through at least one of the membranes when thecartridge is disposed within an associated electric field.
 4. Thecartridge according to claim 1 further comprising two gaskets whereineach gasket is disposed within the interior of the housing and proximateto an outer surface of one of the restriction membranes so as to beadapted to sealingly engage an associated fluid pathway to allow for anelectrically conductive buffer solution to flow along the restrictionmembranes.
 5. The cartridge according to claim 1 wherein the gridelement has a generally planar shape.
 6. The cartridge according toclaim 1 wherein the interior of the grid element is a latticearrangement.
 7. The cartridge according to claim 1 wherein the gridelement is generally elongate in shape and having a first end and asecond end and wherein an aperture is located proximate to each end toallow for the passage of fluid therethrough.
 8. The cartridge accordingto claim 1 wherein at least one of the restriction membranes isgenerally elongate in shape and having a first end and a second end andwherein an aperture is located proximate to each end to allow forpassage of fluid therethrough.